1. Field of the Invention
This invention relates to a method of enhancing growth in patients, particularly those exhibiting a retarded growth rate or weight loss using a combination of natural hormones. In addition, the combination reduces the degree of imbalance of glucose homeostasis induced by either hormone administered alone.
2. Description of Related Art
Insulin-like growth factor I (IGF-I) is a polypeptide naturally occurring in human body fluids, for example, blood and human cerebral spinal fluid. Most tissues and especially the liver produce IGF-I together with specific IGF-binding proteins. These molecules are under the control of growth hormone (GH). Like GH, IGF-I is a potent anabolic protein. See Tanner et al., Acta Endocrinol., 84: 681-696 (1977); Uthne et al., J. Clin. Endocrinol. Metab., 39: 548-554 (1974)). IGF-I has been isolated from human serum and produced recombinantly See, e.g., EP 123,228 and 128,733.
Various biological activities of IGF-I have been identified. Researchers have found that an intravenous bolus injection of IGF-I lowers blood glucose levels in humans. See Guler et al., N. Engl. J. Med., 317: 137-140 (1987). Additionally, IGF-I promotes growth in several metabolic conditions characterized by low IGF-I levels, such as hypophysectomized rats [Guler et al., Endocrinology, 118: Supp 129 abstract, Skottner et al., J. Endocr., 112: 123-132 (1987); Guler et al., Proc. Natl. Acad. Sci. USA, 85: 4889-4893 (1988); Froesch et al., in Endocrinology, Intl. Congress Series 655, ed. by Labrie and Proulx (Amsterdam: Excerpta Medica, 1984), p. 475-479], diabetic rats [Scheiwiller et al., Nature, 323: 169-171 (1986)], and dwarf rats [Skottner et al., Endocrinology. 124: 2519-2526 (1989)]. The kidney weight of hypophysectomized rats increases substantially upon prolonged infusions of IGF-I subcutaneously. Guler et al., Proceedings of the 1st European Congress of Endocrinology, 103: abstract 12-390 (Copenhagen, 1987). The kidneys of Snell dwarf mice and dwarf rats behaved similarly. van Buul-Offers et al., Pediatr. Res., 20: 825-827 (1986); Skottner et al., Endocrinology, supra. An additional use for IGF-I is its administration to improve glomerular filtration and renal plasma flow in human patients. See EP 327,503 published Aug. 9, 1989; Guler et al., Proc. Natl. Acad. Sci. USA, 86: 2868-2872 (1989).
Human growth hormone (hGH) is a single-chain polypeptide consisting of 191 amino acids (molecular weight 2,500). Disulfide bonds link positions 53 and 165 and positions 182 and 189. Niall, Nature, New Biology, 230: 90 (1971). Human GH is a potent anabolic agent, especially due to retention of nitrogen, phosphorus, potassium, and calcium. Treatment of hypophysectomized rats with GH can restore at least a portion of the growth rate of an intact animal Moore et al., Endocrinology, 122: 2920-2926 (1988). Among its most striking effects in hypopituitary (GH-deficient) subjects is accelerated linear growth of bone growth plate cartilage resulting in increased stature. Kaplan, Growth Disorders in Children and Adolescents (Springfield, IL: Charles C. Thomas, 1964).
In 1957, the mechanism of GH action was postulated as being due to GH inducing production of somatomedins (subsequently identified and named IGF-I) in the liver, which travel via the circulation to produce all the effects of GH. Salmon and Daughaday, J. Lab. Clin. Med.. 49: 825-836 (1957). Many studies investigating the relationships among GH, IGF-I, cartilage, cultured human fibroblasts, skeletal muscle, and growth have supported this somatomedin hypothesis. See, e.g., Phillips and Vassilopoulou-Sellin, N. Enol. J. Med., 302: 372-380; 438-446 (1980); Vetter et al., J. Clin. Invest., 7: 1903-1908 (1986); Cook et al., J. Clin. Invest., 81: 206-212 (1988); Isgaard et al., Endocrinology, 123: 2605-2610 (1988); Schoenle et al., Acta Endocrin., 108: 167-174 (1985).
Another theory holds that GH has a direct effect on chondrocytes that is not dependent on circulating IGF-I. For example, several in vivo studies have demonstrated longitudinal long bone growth in rats receiving hGH injected directly into the tibial growth plate [Isaksson et al., Science, 216: 1237-1239 (1982); Russell and Spencer, Endocrinology, 116: 2563-2567 (1985)] or the arterial supply to a limb [Schlechter et al., Am. J. Physiol., 250: E231-235 (1986)]. Additionally it was found that proliferation of cultured lapine ear and rib chondrocytes in culture is stimulated by hGH [Madsen et al., Nature, 304: 545-547 (1983)], this being consistent with a direct GH effect or with an indirect effect of GH mediated by local GH-dependent IGF-I production. Such an autocrine or paracrine model for stimulation of growth has been supported by various lines of experimental evidence. Schlechter et al., Proc. Natl. Acad. Sci. USA. 83: 7932-7934 (1986); Nilsson et al., Calcif. Tissue Int., 40: 91-96 ( 1987). Nilsson et al. showed that while unilateral arterial infusion of IGF-I did not produce a tibial longitudinal bone growth response in hypophysectomized rats, infusion of hGH did induce such growth. Moreover, the influence of GH on the functional maturation of human fetal islet cells in vitro could not be reproduced by adding IGF-I, suggesting a direct rather than a somatomedin-mediated action of GH for these particular cells. Otonkoski et al., Diabetes. 37: 1678-1683 (1988).
A third theory for GH and IGF-I actions is that GH promotes differentiation of stem cells, rendering them responsive to stimulation of proliferation by IGF-I. Green et al., Differentiation, 29: 195-198 (1985). Although support for this model of GH acting to produce IGF-I locally, called the dual effector theory, has been obtained for certain cell types [Zezulak and Green, Science. 233: 551-553 (1986)], its application to skeletal growth has not been established. It has been found that both GH and testosterone could stimulate skeletal growth in the hypophysectomized prepubertal lamb without alteration of circulating IGF-I concentrations, the results not precluding the possibility that the growth-promoting effect of GH was affected by local actions at the site of osteogenesis. Young et al., J. Endocrin., 121: 563-570 (1989). Also, GH has been reported to stimulate tibial epiphyseal plate width in the hypophysectomized rat without increasing circulating IGF-I concentrations. Orlowski and Chernausek, Endocrinol., 123: 44-49 (1988).
More recently, a study was undertaken to reproduce the "direct" in vitro GH effect on epiphyseal and articular chondrocytes to determine whether this effect is mediated by IGF-I in a local autocrine or paracrine fashion. Trippel et al., Pediatr. Res., 25: 76-82 (1989). Human GH was found not to stimulate rabbit articular or epiphyseal chondrocytes or bovine epiphyseal chondrocytes, whereas IGF-I stimulated both mitotic and differentiated cell functions in both epiphyseal and articular chondrocytes. The authors state that the data suggest that the role of IGF-I in skeletal development is complex and may be diverse both in the cellular functions it regulates and the cell populations regulated, requiring further investigation to define the relationship of IGF-I to GH.
It has been reported that the growth response to co-addition of GH and IGF-I was not statistically different from that of GH alone when body weight gain, bone length, or tibial epiphyseal cartilage width was measured. Skottner et al., J. Endocr., supra [iv infusion of bGH (10 mu/day) for 8 days and met-IGF-I (with specific activity of 3400 U/mg, 120 .mu.g/day) for the last 4 days]; Isgaard et al., Am. J. Physiol., 250: E367-E372 (1986) [5 .mu.g of IGF-I and 1 .mu.g of hGH injected locally daily for 5 days]. It was also found that IGF-I, when injected or infused subcutaneously or infused intravenously, is a weak growth promoter in hypophysectomized rats compared with hGH, even when infused in combination with small amounts of hGH. Robinson and Clark, Acta Paediatr. Scand. Supp. 347: 93-103 (1988).
As regards osteoblast-like cells in culture, direct stimulation of their proliferation by hGH is at least partially mediated by IGF-I-like immunoreactivity [Ernst and Froesch, Biochem Biophy Res. Commun., 151: 142-147 (1988)]; the authors found that IGF-I and hGH had additive effects on osteoblast proliferation only when the exogenous IGF-I concentration exceeded that of endogenously produced IGF-I by a large margin. Another in vitro study showed that purified human and synthetic IGF-I stimulated adult articular chondrocyte DNA and proteoglycan synthesis; GH had no effect on either process; and GH added in combination with IGF-I increased proteoglycan, cell-associated proteoglycan, and keratan sulfate synthesis over levels observed with IGF-I alone. Smith et al., J. Orthop. Res., 7: 198-207 (1989). Separate administration of hGH and IGF-I was found to enhance human granulopoiesis, with the effect of hGH on marrow myeloid progenitors apparently mediated by paracrine IGF-I. Merchav et al., J. Clin. Invest., 81: 791-797 (1988). Merchav et al. also noted that myeloid colony formation was significantly enhanced in cultures stimulated with combined limiting concentrations of both IGF-I and hGH, whereas combined maximal concentrations of both peptides did not exert an additive effect.
Also, based on recent immunohistochemical data regarding the GH receptor, it has been suggested that GH may act independently of or synergistically with hepatic IGF-I in carrying out its growth-promoting role in the gastrointestinal tract. Lobie et al., Endocrinol., 126: 299-306 (1990). It has been shown that pretreatment of hypophysectomized rats with GH, but not with IGF-I, promotes the formation of chondrocyte colonies and makes the chondrocytes susceptible to IGF-I in vitro. Lindahl et al., Endocrinol., 121: 1070-1075 (1987). The authors suggest that GH induces colony formation by IGF-I-independent mechanisms and that IGF-I is a second effector in GH action. Further, treatment of hypophysectomized animals with a single dose of hGH restored IGF-I mRNA in parenchymal and in non-parenchymal cells to the extent found in intact animals. van Neste et al., J. Endocr., 119: 69-74 (1988).
However, it has also been reported that IGF-I directly suppresses GH gene transcription and GH secretion at the pituitary level in an inhibitory feedback control mechanism. Namba et al., Endocrinol., 124: 1794-1799 (1989); Yamashita et al., J. Biol. Chem., 262: 13254-3257 (1987). Additionally, it was reported that the maximum stimulation of glucose metabolism in 3T3 adipocytes achieved by hGH is only a fraction of that produced by various IGFs, indicating that extracellular IGFs do not mimic the effects of hGH on glucose metabolism in these adipocytes. Schwartz et al., Proc. Natl. Acad. Sci. USA, 82: 8724-8728 (1985). Moreover, human GH was found not to enhance further the IGF-I-stimulated Leydig cell steroidogenesis. Horikawa et al., Eur. J. Pharmacol., 166: 87-94 (1989). Another negative finding was that the combination of chick growth hormone and human IGF-I did not stimulate cell proliferation and metabolic activity of cultured epiphyseal growth plate chondrocytes above human IGF-I alone. Rosselot et al., The Endocrine Society 72nd Annual Meeting, abstract 202, p. 75, of Program and Abstracts released prior to the meeting in Atlanta, GA on June 20-23, 1990. It has also been reported that both hGH and hIGF-I can promote growth in the mutant dwarf rat, but they differ both quantitatively and qualitatively in their pattern of actions. Skottner et al., Endocrinology, supra. Additionally, a loss of IGF-I receptors in cultured bovine articular chondrocytes was found after pre-exposure of the cells to pharmacological doses of either hGH or bGH. Watanabe et al., J. Endocr., 107: 275-283 (1985). The necessity for large amounts of GH is attributed to extremely low affinity of GH binding sites on these cells. The authors speculate that living organisms have a protection mechanism to avoid unnecessary overgrowth of the body resulting in down-regulation of the IGF-I receptors.
U.S. Pat. No. 4,857,505 issued Aug. 15, 1989 discloses use of an adduct of a growth hormone, growth factor, IGF-I, or fragment thereof covalently bonded to an activated polysaccharide for increased half-life, increased weight gain in animals, and increased milk production.
Known side effects of hGH treatment include sodium retention and expansion of extracellular volume [Ikkos et al., Acta Endocrinol. (Copenhagen), 32: 341-361 (1959); Biglieri et al., J. Clin. Endocrinol. Metab., 21: 361-370 (1961)], as well as hyperinsulinemia and hyperglycemia. The major apparent side effect of IGF-I is hypoglycemia. Guler et al., Proc. Natl. Acad. Sci. USA, 1989, supra.
It is an object of the present invention to provide a combination of IGF-I and GH that has a greater effect on growth of patients than either hormone alone.
It is another object to provide a method for treating patients, whether children or adults, that experience a reduced anabolic effect of GH, as by a reduced ability to produce an IGF-I response to the GH, or experience diabetogenic effects or other side effects when treated with GH alone, or hypoglycemia when treated with IGF-I alone.
These and other objects will be apparent to those of ordinary skill in the art.